Establishment of a functional secretory IgA transcytosis model system in vitro for functional food screening

Appl Microbiol Biotechnol. 2015 Jul;99(13):5535-45. doi: 10.1007/s00253-015-6501-9. Epub 2015 Mar 18.

Abstract

A characteristic mucosal immune response involves the production of antigen-specific secretory immunoglobulin A (SIgA) antibodies. In order to study transcytosis by mimicking the SIgA secretion and to screen for SIgA secretion-promoting substances, we developed a model system of a transfected Madin-Darby canine kidney (MDCK) cell line that expresses the human polymeric immunoglobulin receptor (pIgR). We thus isolated the human dIgA (dimeric IgA)/pIgA (polymeric IgA) complex as the binding ligands. In the present study, a recombinant vector encoding the human pIgR gene was constructed and infected into MDCK cells. Following reverse transcription polymerase chain reaction (RT-PCR), immunoblotting, and immunofluorescence staining, we confirmed that pIgR was expressed in the transfectant MDCK-pIgR cells and was located at the basolateral side of the cell surface. We also confirmed the coexistence of the dIgA/pIgA complex in the IgA myeloma serum. The covalent dIgA/pIgA complex was then isolated from the serum of an IgA multiple myeloma patient using an ÄKTA purifier operation system with a HiPrep 16/60 Sephacryl S-300 HR column, in order to utilize the complex as transcytosis ligands for human pIgR. Finally, we confirmed the uptake of the isolated human dIgA/pIgA complex into MDCK-pIgR cells. We demonstrated that the human dIgA/pIgA complex was transcytosed into the apical side of the monolayer cells. Therefore, our MDCK-human pIgR cell transcytosis model is an operational system and can be used for screening functional food components that promote dIgA/pIgR transcytosis as well as SIgA secretion.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Dogs
  • Fluorescent Antibody Technique
  • Functional Food*
  • Humans
  • Immunoblotting
  • Immunoglobulin A / blood
  • Immunoglobulin A, Secretory / metabolism*
  • Madin Darby Canine Kidney Cells
  • Mass Screening / methods*
  • Models, Biological
  • Multiple Myeloma / immunology
  • Receptors, Polymeric Immunoglobulin / genetics
  • Receptors, Polymeric Immunoglobulin / metabolism
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Reverse Transcriptase Polymerase Chain Reaction
  • Serum / immunology
  • Transcytosis*
  • Transgenes

Substances

  • Immunoglobulin A
  • Immunoglobulin A, Secretory
  • Receptors, Polymeric Immunoglobulin
  • Recombinant Proteins
  • polymeric IgA